Natural killer (NK) cells are lymphocytes of innate immunity that secrete cytokines and kill cells at early times of infection. Besides defending against infection, NK cells also control trophoblast invasion during implantation in pregnancy and can mediate graft-versus-leukemia responses following allogeneic bone-marrow transplantation. NK-cell responses are controlled by systems of activating and inhibitory receptors, some of which recognize MHC class I or class l-like ligands. Of these, the killer-cell immunoglobulin-like receptors (KIR) are highly diverse, rapidly evolving and recognize polymorphic HLA class I molecules. Although vestigial in mice, the KIR system is highly elaborated in the human species. Notably, the telomeric part of the KIR locus, which contains the highly polymorphic KIR3DL1 and KIR3DL2 genes, is expanded in humans compared to other primate species. KIR3DL1 encodes inhibitory receptors specific for HLA-B determinants. Allotypes of KIR3DL1 are distinguished by their levels of expression at the cell surface, a modulation over an order of magnitude that is proposed to be a main function of the polymorphism. Three aims will work together to test this hypothesis. In aim 1, the role of protein polymorphism in setting cell surface levels will be investigated by comparison of naturally occurring allotypes and mutants in which the naturally occurring substitutions are swapped. Analysis will use a newly developed assay for quantifying the relative amount of cellular KIR3DL1 that is present on the cell surface. Aim 2 will investigate the influence of promoter and other non-coding polymorphisms of the KIR3DL1 gene upon the surface level of KIR3DL1 expression. Thus aims 1 and 2 will define the precise mechanisms that set the level of surface expression for each of fourteen KIR3DL1 allotypes. Aim 3 will investigate the effects on NK-cell functions of changing the levels of KIR3DL1 expression in the absence of change in the protein sequence. This will separate other potential effects of the polymorphism, such as ligand specificity, from the level of cell-surface expression.